An IS4 insertion at the glnA control region of Escherichia coli creates a new promoter by providing the -35 region of its 3'-end.
An insertion element (IS)4 insertion selected as suppressor of the rpoN73::Tn5 alelle was located inside the control region of the glnA gene in Escherichia coli. In the rpoN73::Tn5 background the IS4 insertion promotes glnA transcription at a low constitutive level sufficient to sustain glutamine-independent growth. The IS4 insertion mutation in either rpoN73::Tn5 or wild-type backgrounds promotes glnA transcription from a new start site located two bases downstream of the glnAp2 start site. Analysis of sequences flanking the insertion point showed a promoter sequence whose -35 region was located inside the IS4 sequence and the -10 region was inside the glnA control region. Site-directed mutagenesis of relevant nucleotide residues of the newly created promoter impaired transcription of a reporter gene. The results support our contention that IS4 carries a -35 promoter region that is able to create functional hybrid promoters. We propose that this mechanism could be one of the molecular reasons of the suppressor activity previously reported for IS4.
Camarena
L
L
Departamento de Biología Molecular, Instituto de Investigaciones Biomédicas, U.N.A.M., México, D.F. rosal@servidor.unam.mx
Poggio
S
S
Campos
A
A
Bastarrachea
F
F
Osorio
A
A
D004251:DNA Transposable Elements; D004268:DNA-Binding Proteins; D012321:DNA-Directed RNA Polymerases / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins; D015964:Gene Expression Regulation, Bacterial; D005974:Glutamate-Ammonia Ligase / Q000235:genetics* / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D011401:Promoter Regions, Genetic; D051877:RNA Polymerase Sigma 54; D012808:Sigma Factor / Q000235:genetics / Q000378:metabolism
D004251:DNA Transposable Elements; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D012808:Sigma Factor; C497751:rpoN protein, E coli; D012321:DNA-Directed RNA Polymerases; D051877:RNA Polymerase Sigma 54; C081173:glutamine synthetase I; D005974:Glutamate-Ammonia Ligase